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Data associated with 'An in vivo platform to select and evolve aggregation-resistant proteins'
Citation
Ebo, Jessica S. and Saunders, Janet C. and Devine, Paul W. A. and Gordon, Alice M. and Warwick, Amy S. and Schiffrin, Bob and Chin, Stacey E. and England, Elizabeth and Button, James D. and Lloyd, Christopher and Bond, Nicholas J. and Ashcroft, Alison E. and Radford, Sheena E. and Lowe, David and Brockwell, David J., AstraZeneca (2019) Data associated with 'An in vivo platform to select and evolve aggregation-resistant proteins'. University of Leeds. [Dataset] https://doi.org/10.5518/739
Dataset description
Protein biopharmaceuticals are highly successful, but their utility is compromised by their propensity to aggregate during manufacture and storage. As aggregation can be triggered by non-native states, whose population is not necessarily related to thermodynamic stability, prediction of poorly-behaving biologics is difficult, and searching for sequences with desired properties is labour-intensive and time-consuming. Here we show that an assay in the periplasm of E. coli linking aggregation directly to antibiotic resistance acts as a sensor for the innate (un-accelerated) aggregation of antibody fragments. Using this assay as a directed evolution screen, we demonstrate the generation of aggregation resistant scFv sequences when reformatted as IgGs. This powerful tool can thus screen and evolve ‘manufacturable’ biopharmaceuticals early in industrial development. By comparing the mutational profiles of three different immunoglobulin scaffolds, we show the applicability of this method to investigate protein aggregation mechanisms important to both industrial manufacture and amyloid disease.
| Keywords: | Protein aggregation, biopharmaceuticals, directed evolution | ||||||
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| Subjects: | C000 - Biological sciences > C700 - Molecular biology, biophysics & biochemistry | ||||||
| Divisions: | Faculty of Biological Sciences > Astbury Centre for Structural Molecular Biology Faculty of Biological Sciences > School of Molecular and Cellular Biology |
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| License: | Creative Commons Attribution 4.0 International (CC BY 4.0) | ||||||
| Date deposited: | 03 Dec 2019 14:32 | ||||||
| URI: | https://archive.researchdata.leeds.ac.uk/id/eprint/610 | ||||||
Files
Data
In vivo growth of biopharmaceutical scaffolds [16kB]  |
Comparison of point mutations on WFL growth [10kB] |
Comparison of point mutations on WFL SEC retention time [8kB] |
Evolution of WFL in vivo [11kB] |
Evolved WFL in vitro [8kB] |
Evolved IgGs IC50 [12kB] |
WFL Mutational hotspots [15kB] |
Li33 Mutational hotspots [14kB] |
JTO Mutational hotspots [11kB] |
Computational aggregation hotspots [20kB] |
Thermal stability of WFL IgGs [119kB] |
In vivo growth of evolved Li33 variants [9kB] |
In vivo growth of evolved JTO variants [9kB] |
Thermal stability of Li33 IgGs [47kB] |
Li33 AC-SINS [8kB] |
Li33 PEG solubility [10kB] |
In vivo growth of WFL hotspot residues [11kB] |
In vivo growth of biopharmaceutical scaffolds [16kB] 
In vivo growth of biopharmaceutical scaffolds [16kB]